Scientists Target Incurable Mitochondrial Diseases With New Gene Editing Tools

The energy factories in our cells contain their own genes, and genetic mutations in them can cause deadly inherited diseases.

These oblong-shaped organelles, or mitochondria, translate genes into proteins, which together form a kind of production chain that supplies cells with energy. Mutations in mitochondrial DNA, or mtDNA, torpedo the process, leading to sluggish cells that eventually wither away.

Some mitochondrial DNA mutations have been linked to age-related diseases, metabolic problems, and stroke-like symptoms. Others are involved in epilepsy, eye diseases, cancer, and cognitive troubles. Many of the diseases are inherited. But none are treatable.

“Mitochondrial disorders are incredibly diverse in their manifestation and progression… [and] therapeutic options for these pathologies are rarely available and only moderately effective,” wrote Alessandro Bitto at the University of Washington last year.

As a workaround, some countries have already approved mitochondrial transfer therapy, which replaces defective mitochondria with healthy ones in reproductive cells. The resulting “three-parent” kids are generally healthy. But the procedure remains controversial because it involves tinkering with human reproductive cells, with potentially unknown repercussions down the line.

The new study, published in Science Translational Medicine, took an alternative approach—gene therapy. Using a genetic tool called base editing to target mitochondrial DNA, the team successfully rewrote damaged sections to overcome deadly mutations in mice.

“This approach could be potentially used to treat human diseases,” wrote the team.

Double Trouble

Our genetic blueprints are housed in two places. The main set is inside the nucleus. But there’s another set in our mitochondria, the organelles that produce over 90 percent of a cell’s energy.

These pill-shaped structures are enveloped in two membranes. The outer membrane is structural. The inner membrane is like an energy factory, containing teams of protein “workers” strategically placed to convert food and oxygen into fuel.

Mitochondria are strange creatures. According to the latest theory, they were once independent critters that sheltered inside larger cells on early Earth. Eventually, the two merged into one. Mitochondria offered protocells a more efficient way to generate energy in exchange for safe haven. Eventually, the team-up led to all the modern cells that make up our bodies.

This is likely why mitochondria have their own DNA. Though it’s separate, it works the same way: Genes are translated into messenger RNA and shuttled to the mitochondria’s own protein-making factories. These local factories recruit “transporters,” or mitochondrial transfer RNA, to supply protein building blocks, which are stitched into the final protein product.

These processes happen in solitude. In a way, mitochondria reign their own territory inside each cell. But their DNA has a disadvantage. Compared to our central genetic blueprint, it’s more prone to mutations because mitochondria evolved fewer DNA repair abilities.

“There are about 1,000 copies of mtDNA in most cells,” but mutations can coexist with healthy variants, the authors wrote. Mitochondrial diseases only happen when mutations overrun healthy DNA. Even a small amount of normal mitochondrial DNA can protect against mutations, suggesting gene editing could be a way to tackle these diseases.

Into the Unknown

Current treatments for people with mitochondrial mutations ease symptoms but don’t tackle the root cause.

One potential therapy under development would help cells destroy damaged mitochondria. Here, scientists design “scissors” that snip mutated mitochondrial DNA in cells also containing healthy copies. By cutting away damaged DNA, it’s hoped healthy mitochondria repopulate and regain their role.

In 2020, a team led by David Liu at MIT and Harvard’s Broad Institute of MIT and Harvard unleashed a gene editing tool tailored to mitochondria. Well-known for his role in developing CRISPR base editing—a precision tool to swap one genetic letter for another—his lab’s tool targeted mitochondrial DNA with another method.

They broke a bacterial toxin into two halves—both are inactive and non-toxic until they join together at a targeted DNA site. When activated, the editor turns the DNA letter “C” to “T” inside mitochondria, with minimal changes to other genetic material.

In the new study, the team focused on a mitochondrial defect that damages the organelles’ “transporter” molecules. Without this transfer RNA, mitochondria can’t make the proteins that are essential for generating energy.

The transporter molecules look like four-leaf clovers with sturdy stems. Each leaf is made of a pair of genetic letters that grab onto each other. But in some mutations, pairs can’t hook together, so the leaves no longer connect, and they wreck the transporter’s function.

Powering Up

These early results suggest that DNA mutations in mitochondria damage the cell’s ability to provide energy. Correcting the mutations may help.

As a test, the team used their tool to transform genetic letters in cultured cells. After several rounds of treatment, 75 percent of the cells had reprogrammed mitochondria.

The team then combined the editor with a safe delivery virus. When injected into the bloodstreams of young adult mice, the editor rapidly reached cells in their hearts and muscles. In hearts, the treatment upped normal transfer RNA levels by 50 percent.

It’s not a perfect fix though. The injection didn’t reach the brain or kidneys, and they found very few signs of editing in the liver. This is surprising, wrote the authors, because the liver is usually the first organ to absorb gene editors.

When the team upped the dose, off-target edits in healthy mitochondria skyrocketed. On the plus side, the edits didn’t notably alter the main genetic blueprints contained in nuclear DNA.

It’ll be a while before mitochondrial gene editors can be tested in humans. The current system uses TALE, an older gene editing method that’s regained some steam. Off-target edits, especially at higher doses, could also potentially cause problems in unexpected tissues or organs.

“Specific tissues may respond differently to editing, so optimization should also consider the possibility of the target tissue being more sensitive to undesirable off-target changes,” wrote the team.

Overall, there’s more work to do. But new mitochondrial base editors “should help improve the precision of mitochondrial gene therapy,” the team wrote.